Beyond the “Recognition Code” Scot A. Wolfe, Robert A. Grant, Monicia Elrod-Erickson, Carl O. Pabo  Structure  Volume 9, Issue 8, Pages 717-723 (August 2001) DOI: 10.1016/S0969-2126(01)00632-3

Figure 1 Comparison of the DNA Recognition of the TATAZF, TATAZF*, and Zif268 Fingers (a) Left panel: structure of TATAZF bound to a 16 bp oligonucleotide. At this level of detail, this structure is virtually indistinguishable from the TATAZF*-DNA complex. Right panel: superposition of the TATAZF and Zif268 structures [4] (protein: red and gray, respectively; DNA: blue and magenta, respectively). Each protein uses a tandem array of three fingers to recognize its binding site. Every finger is folded into a ββα motif around a single metal ion (displayed as a van der Waals surface). The α helix of each finger fits into the major groove, and base-specific recognition is mediated by residues within (or immediately preceding) this helix. (b) Aligned sequence of the three Cys2His2 zinc fingers from Zif268 [26], with the residues involved in base contacts highlighted in blue [3, 4]. The α-helical region of each finger is denoted by a cylinder (above), and β strands are indicated by filled arrows. The numbers just below the orange cylinder indicate the position of the residue within the recognition helix, with −1 indicating the residue immediately preceding the helix. (c) Sequences of the recognition helices from Zif268, TATAZF, and TATAZF* displaying all of the positions (−1, 1, 2, 3, 5, and 6) that had been randomized by Greisman and Pabo [7]. Residues involved in base contacts are shown in blue. Residues involved in interfinger interactions are indicated in red. (d) Diagram comparing the base contacts made by Zif268, TATAZF, and TATAZF*. Hydrophobic or van der Waals interactions are indicated by black dots, and hydrogen bonds are indicated by arrows. Color coding of the DNA region contacted by each finger emphasizes the size (and overlapping arrangement) of the binding sites. Red bars between residues at positions 6 and −1 on neighboring fingers indicate the presence of interfinger interactions. (e) Diagram comparing the DNA recognition pattern of the fingers from Zif268 and the TATA complexes. Solid lines denote DNA interactions that occur in most of the fingers, while dashed lines indicate DNA interactions that occur less frequently Structure 2001 9, 717-723DOI: (10.1016/S0969-2126(01)00632-3)

Figure 2 Unexpected DNA Contacts Are Observed from Position 1 of the Recognition Helix In fingers 2 and 3 of both TATA complexes, the amino acid at position 1 interacts with the secondary strand of the DNA as well as with the sugar-phosphate backbone. In this example, Leu-75 in finger 3 of TATAZF makes contacts to bases T7* and A6* on the secondary strand of the DNA. (Note: In this and subsequent figures, interactions of special interest are indicated by dotted lines. Numbers near dotted lines indicate the distance in angstroms. Zinc fingers are modeled as ribbons that are color coded by finger as shown in Figure 1d. The primary DNA strand is colored brown, and the secondary stand is colored magenta. Asterisks denote DNA bases in the secondary strand. Portions of Figure 1d that correspond to the displayed region of the structure are shown as an inset for each of the more detailed pictures.) Structure 2001 9, 717-723DOI: (10.1016/S0969-2126(01)00632-3)

Figure 3 Interfinger Interactions Play Important Roles in Stabilizing the Protein-DNA Interface (a) In TATAZF, Thr-24 (position 6 of finger 1) interacts with the side chain of Gln-46 (position −1 of finger 2) which in turn packs against the methyl group of T6. Presumably, these interactions stabilize the DNA contact from Gln-46 to A5. Thr-24 also interacts with Arg-27 to further organize the interfinger junction. Arg-27, in turn, makes interfinger contacts to Pro-34 and to the backbone carbonyl of Ser-45. (b) Although they have received no attention in previous studies, interfinger interactions also occur in the Zif268 structure [4]. As illustrated here, both Thr-52 (position 6 of finger 2) and Asp-76 (position 2 of finger 3) help to stabilize the interaction of Arg-74 (position −1 of finger 3) with a guanine on the primary strand of the DNA. Interfinger interactions between position 6 of one finger and position −1 of a neighboring finger also occur in the GLI structure [27]. (Coloring scheme is the same as in Figure 2.) Structure 2001 9, 717-723DOI: (10.1016/S0969-2126(01)00632-3)

Figure 3 Interfinger Interactions Play Important Roles in Stabilizing the Protein-DNA Interface (a) In TATAZF, Thr-24 (position 6 of finger 1) interacts with the side chain of Gln-46 (position −1 of finger 2) which in turn packs against the methyl group of T6. Presumably, these interactions stabilize the DNA contact from Gln-46 to A5. Thr-24 also interacts with Arg-27 to further organize the interfinger junction. Arg-27, in turn, makes interfinger contacts to Pro-34 and to the backbone carbonyl of Ser-45. (b) Although they have received no attention in previous studies, interfinger interactions also occur in the Zif268 structure [4]. As illustrated here, both Thr-52 (position 6 of finger 2) and Asp-76 (position 2 of finger 3) help to stabilize the interaction of Arg-74 (position −1 of finger 3) with a guanine on the primary strand of the DNA. Interfinger interactions between position 6 of one finger and position −1 of a neighboring finger also occur in the GLI structure [27]. (Coloring scheme is the same as in Figure 2.) Structure 2001 9, 717-723DOI: (10.1016/S0969-2126(01)00632-3)

Figure 4 Side Chain-Side Chain Interactions within the Recognition Helix Stabilize the Protein-DNA Interface In the selected TATA proteins, whenever glutamine was present at position 6 of finger 2, threonine was absolutely conserved at position 2 [7]. The structure of TATAZF* explains this preference; the γ-methyl group of Thr-48 and the methyl group of T8 pack against opposite faces of Gln-52. Presumably, this stabilizes the interaction of Gln-52 with A7. This is analogous to the interfinger interaction described between fingers 1 and 2 of TATAZF in Figure 3a. (Coloring scheme is the same as in Figure 2.) Structure 2001 9, 717-723DOI: (10.1016/S0969-2126(01)00632-3)